Adhesion between a cell and the extracellular matrix is mediated by a transmembrane cell adhesion protein as typified by integrins. Integrins are heterodimers that have 1:1 of α- and β-chains. So far, 18 types of α-chains and 8 types of β-chains have been found where at least 24 types of combinations thereof have been identified and confirmed. Each integrin is known to recognize a specific extracellular matrix (ligand). Moreover, the transmembrane cell adhesion protein containing an integrin not only plays a role of adhering and anchoring a cell and the extracellular matrix but it has also been found to convert information from the extracellular matrix into intracellular signals for regulating cell proliferation, motility, cell death, differentiation and the like.
Integrins are classified, according to their specificities and functions with respect to ligands, into subfamilies, namely, collagen receptors, laminin receptors, RGD receptors that recognize Arg-Gly-Asp (RGD) sequence contained in fibronectin, vitronectin and the like, and leukocyte-specific receptors that exist only in leukocytes. Alpha-4 integrins and α9 integrins do not belong to any of these subfamilies and are called α4 integrin subfamily.
Ligands that are known to bind to α4 and α9 integrins include osteopontin (hereinafter, referred to as OPN), EDA domain of fibronectin, propeptide-von Willebrand factor (pp-vWF), tissue transglutaminase (tTG), coagulation factor XIII and Vascular Cell Adhesion Molecule-1 (VCAM-1). Furthermore, ligands that are known to be recognized specifically by α4 integrin include CS-1 domain of fibronectin, MadCAM-1 (α4β7) and the like. Meanwhile, ligands that are known to be recognized specifically by α9 integrin include tenascin C, plasmin and the like.
OPN, one type of extracellular matrices (ECM), is a secreted acid phosphorylated acid glycoprotein with a molecular weight of about 41 kDa, which is a molecule generally acknowledged to be expressed in breast milk, urine, renal tubules, osteoclasts, osteoblasts, macrophage, activated T cell, tumor tissue and the like. OPN has cell adhesion sequence GRGDS (SEQ ID NO:17) in the middle, and SVVYGLR (SEQ ID NO:18) or SLAYGLR (SEQ ID NO:19) sequence for human or murine OPN, respectively, which is immediately followed by a thrombin cleavage site. OPN adheres to integrin of RGD receptor via the GRGDS sequence (SEQ ID NO:17) and to α4 (α4β1) and α9 (α9β1) integrins via the SVVYGLR sequence (SEQ ID NO:18) or the SLAYGLR sequence (SEQ ID NO:19).
While α4β1 binds to OPN that is not cleaved with thrombin (non-cleaved OPN) as well as an N-terminal fragment cleaved with thrombin (cleaved OPN), α9β1 differs in that it binds only to cleaved OPN.
The amino acid sequences of α4,α9 and β1 integrin subunits are known and are registered with GenBank. The amino acid sequences are known to be highly similar among the species.
WO02/081522 discloses a therapeutic effect for rheumatoid arthritis and hepatitis by utilizing an OPN-deficient mouse and a neutralizing antibody against OPN to suppress the OPN functions. This publication also discloses that the SVVYGLR sequence (SEQ ID NO:18), i.e., a sequence that recognizes α4 and α9 integrins, plays an important role upon onset of an inflammatory disease and that a receptor for OPN involved in inflammatory diseases is expressed in the immunocompetent cell or the like.